Interferons (IFNs) are key cytokines in the establishment of a multifaceted antiviral response. Based on the structural features, receptor usage and biological activities, there are (3) distinct types of IFNs, commonly recognized as type I, II, and III IFNs.
Although IFNs are important mediators of antiviral protection, their roles in antiviral defense vary. For example, type I IFNs (IFN-α/β/ω/εκ in humans) possess strong intrinsic antiviral activity, and are able to induce a potent antiviral state in a wide variety of cells (Levy and Garcia-Sastre 2001). In contrast, studies with IFN-γ and IFN-γ receptor knock-out mice (Dalton et al., 1993) as well as analysis of humans who possess inherited genetic mutations of the IFN-γ receptor (Dorman et al., 2004) reveal that antiviral activity is not the primary biological function of IFN-γ. IFN-γ is classified as a Th1-type cytokine that stimulates cell-mediated immune responses that are critical for the development of host protection against pathogenic intracellular microorganisms such as Mycobacterium tuberculosis (Bach et al., 1997) as well as antitumor immune responses.
The most recent additions are the type III IFNs (IFN-λs). They demonstrate structural features of the IL-10-related cytokines but also induce antiviral activity in a variety of target cells, which supports their functional classification as a new type of IFNs (Kotenko et al., 2003). The Interferon-λ (IFN-λ) family was reported in early 2003 (Kotenko et al., 2003; Sheppard et al., 2003). Three (3) IFN-λ genes were identified as encoding three (3) distinct, albeit highly-related, proteins denoted as IFN-λ1, IFN-λ2, and IFN-λ3, respectively. Together, they comprise the type III subset of interferons (IFNs)—distinct from both type I and type II IFNs. Phylogenetically, the IFN-λ genes reside somewhere between the type I IFN and IL-10 gene families. Amino acid sequence comparisons show that the type III IFNs exhibit about ˜5%-18% identity with either type I IFNs or the IL-10-related cytokines.
Direct detection techniques for IFN-λ2 or IFN-λ3 mRNA include PCR techniques involving a design of primers and probe-based hybridization. However, the mRNA sequences of IFN-λ2 and IFN-λ3 in human exhibit a 98% identity. Given such a high degree of identity, it is commonly known to be difficult to distinguish these two (2) mRNAs by routine molecular assays such as qRT-PCR. Put simply, it is difficult to identify unique primer pairs that are specific for IFN-λ2 mRNA but not IFN-λ3 mRNA, and vice-versa. To the best of the inventors' knowledge, there is no PCR techniques that can identify and distinguish human IFN-λ2 or IFN-λ3 mRNAs.
There is a continuing need to develop a molecular diagnostic assay that can specifically detect human IFN-λ2 mRNA but not IFN-λ3 mRNA or vice versa.